Benzocyanine compounds

Patent metadata

Field	Value
Publication number	US-9751868-B2
Application number	US-201313778678-A
Country	US
Kind code	B2
Filing date	Feb 27, 2013
Priority date	Feb 28, 2012
Publication date	Sep 5, 2017
Grant date	Sep 5, 2017

Abstract

Compounds useful as labels with properties comparable to known fluorescent compounds. The compounds are conjugated to proteins and nucleic acids for biological imaging and analysis. Synthesis of the compounds, formation and use of the conjugated compounds, and specific non-limiting examples of each are provided.

First claim

What is claimed is:

1. A compound selected from the group consisting of

where
each of R 1 , R 2 , R 5 , and R 6 is the same or different and is independently selected from the group consisting of an aliphatic, heteroaliphatic, sulfoalkyl, heteroaliphatic with terminal SO 3 , a PEG group P—Z where P is selected from an ethylene glycol group, a diethylene glycol group, and a (poly)ethylene glycol group, where the (poly)ethylene glycol group is (CH 2 CH 2 O) s , where s is an integer from 3-6 inclusive, a sulfonamide group -L-SO 2 NH—P—Z, and a caboxamide group -L-CONH—P—Z, where Z is selected from H, CH 3 , a CH 3 group, an alkyl group, or a heteroalkyl group;
each of R 7 , R 8 , R 11 , R 12 , R 13 , and R 14 is the same or different and is independently selected from the group consisting of H, SO 3 , a PEG group P—Z where P is selected from an ethylene glycol group, a diethylene glycol group, and a (poly)ethylene glycol group, where the (poly)ethylene glycol group is (CH 2 CH 2 O) s , where s is an integer from 3-6 inclusive, a sulfonamide-containing group -L-SO 2 NH—P—Z, and a carboxamide-containing group -L-CONH—P—Z, where Z is selected from H, CH 3 , a CH 3 group, an alkyl group, or a heteroalkyl group;
L is selected from a divalent linear (—(CH 2 ) t —, t=0 to 15), crossed, or cyclic alkyl group optionally substituted by at least one oxygen atom and/or sulfur atom;
Kat is a number of Na + , K + , Ca 2+ , ammonia, or other cation(s) needed to compensate the negative charge brought by the cyanine; m is an integer from 0 to 5 inclusive; p is an integer from 1 to 6 inclusive;
each of R3 and R4 is the same or different and is independently hydrogen, an aliphatic group, a heteroaliphatic group, or a PEG group P—Z where P is selected from an ethylene glycol group, a diethylene glycol group, and a (poly)ethylene glycol group, where the (poly)ethylene glycol group is (CH 2 CH 2 O) s , where s is an integer from 3-6 inclusive, and Z is selected from H, a CH 3 group, an alkyl group, or a heteroalkyl group; or R3 and R4 together form a cyclic structure where R3 and R4 are joined using a divalent structural element selected from the group consisting of —(CH 2 ) q —, —(CH 2 ) q O(CH 2 ) q′ —, —(CH 2 ) q S(CH 2 ) q′ —, —(CH 2 ) q CH═CH—, —OCH═CH— where each of q and q′ is the same or different and is a integer from 2 to 6 inclusive; and
Y is selected from the group consisting of hydrogen, alkyl, sulfoalkyl, fluorine, chlorine, bromine, a substituted or unsubstituted aryl-, phenoxy-, phenylmercapto function, a PEG group P—Z where P is selected from an ethylene glycol group, a diethylene glycol group, and a (poly)ethylene glycol group, where the (poly)ethylene glycol group is (CH 2 CH 2 O) s , where s is an integer from 3-6 inclusive, and Z is selected from H, a CH 3 group, an alkyl group, or a heteroalkyl group, and an oxygen-containing group OR PM , where R PM is selected from the group consisting of hydrogen, a substituted or unsubstituted alkyl group, a substituted or unsubstituted heteroalkyl group, a substituted or unsubstituted cyclic alkyl group, a substituted or unsubstituted heterocyclic alkyl group, a substituted or unsubstituted aryl group, and a substituted or unsubstituted heteroaryl group, where the group can be substituted one or more times with one or more of the following groups: hydroxyl, sulfo, carboxy, and/or amino.

2. The compound of claim 1 wherein each of R13 and R14 is sulfo.

3. The compound of claim 1 wherein each of R7, R8, R11, and R12 is sulfo.

4. A method of labeling at least one biomolecule, the method comprising combining a composition comprising at least one excipient and the compound of claim 1 in an effective concentration with at least one biomolecule under conditions sufficient for labeling the biomolecule with the compound.

5. The method of claim 4 wherein the biomolecule is selected from the group consisting of a protein, antibody, enzyme, nucleoside triphosphate, oligonucleotide, biotin, hapten, cofactor, lectin, antibody binding protein, carotenoid, carbohydrate, hormone, neurotransmitter, growth factors, toxin, biological cell, lipid, receptor binding drug, fluorescent proteins, organic polymer carrier material, inorganic polymeric carrier material, and combinations thereof.

6. A method of detecting at least one biomolecule, the method comprising combining a composition comprising at least one excipient and the compound of claim 1 in an effective concentration with at least one biomolecule under conditions sufficient for binding the compound to the biomolecule, and detecting the biomolecule-bound compound.

7. The method of claim 6 wherein the biomolecule is selected from a protein, antibody, enzyme, nucleoside triphosphate, oligonucleotide, biotin, hapten, cofactor, lectin, antibody binding protein, carotenoid, carbohydrate, hormone, neurotransmitter, growth factors, toxin, biological cell, lipid, receptor binding drug, fluorescent proteins, organic polymer carrier material, inorganic polymeric carrier material, and combinations thereof.

8. The method of claim 6 wherein the at least one biomolecule is detected in an assay selected from fluorescence microscopy, flow cytometry, in vivo imaging, immunoassay, hybridization, chromatographic assay, electrophoretic assay, microwell plate based assay, fluorescence resonance energy transfer (FRET) system, bioluminescence reasonance energy transfer (BRET), high throughput screening, or microarray.

9. The method of claim 6 wherein the biomolecule is detected by in vivo imaging comprising providing the biomolecule-bound compound to at least one of a biological sample, tissue, or organism, and detecting the biomolecule within the at least one of a biological sample, tissue, or organism.

10. A kit for detecting and/or labeling at least one biomolecule in a sample, the kit comprising the compound of claim 1 and at least one excipient, and instructions for use of the compound to detect a biomolecule in a sample.

11. A compound selected from the group consisting of

where
each of R 1 , R 2 , R 5 , and R 6 is the same or different and is independently selected from the group consisting of an aliphatic, heteroaliphatic, sulfoalkyl, heteroaliphatic with terminal SO 3 , a PEG group P—Z where P is selected from an ethylene glycol group, a diethylene glycol group, and a (poly)ethylene glycol group, where the (poly)ethylene glycol group is (CH 2 CH 2 O) s , where s is an integer from 3-6 inclusive, a sulfonamide group -L-SO 2 NH—P—Z, and a caboxamide group -L-CONH—P—Z, where Z is selected from H, CH 3 , a CH 3 group, an alkyl group, or a heteroalkyl group;
each of R 7 , R 8 , R 11 , R 12 , R 13 , and R 14 is the same or different and is independently selected from the group consisting of H, SO 3 , a PEG group P—Z where P is selected from an ethylene glycol group, a diethylene glycol group, and a (poly)ethylene glycol group, where the (poly)ethylene glycol group is (CH 2 CH 2 O) s , where s is an integer from 3-6 inclusive, a sulfonamide-containing group -L-SO 2 NH—P—Z, and a carboxamide-containing group -L-CONH—P—Z, where Z is selected from H, CH 3 , a CH 3 group, an alkyl group, or a heteroalkyl group;
X is selected from —OH, —SH, —NH 2 , —NH—NH 2 , —F, —Cl, —Br, —I, —NHS (hydroxysuccinimidyl/sulfosuccinimidyl), —O-TFP (2,3,5,6-tetrafluorophenoxy), —O-STP (4-sulfo-2,3,5,6-tetrafluorophenoxy), —O-benzotriazole, -benzotriazole, imidazole, azide, —O-carbodiimide, —NR-L-OH, —NR-L-O-phosphoramidite, —NR-L-SH, —NR-L-NH 2 , —

Assignees

• Hermanson Greg
• Czerney Peter T
• Desai Surbhi
• Wenzel Matthias S
• Dworecki Boguslawa R
• Lehmann Frank G
• Nlend Marie Christine
• Pierce Biotechnology Inc
• Dyomics Gmbh

Inventors

• Hermanson Greg
• Czerney Peter T
• Desai Surbhi
• Wenzel Matthias S
• Dworecki Boguslawa R
• Lehmann Frank G
• Nlend Marie Christine

Classifications

• G01N33/54346

  Nanoparticles · CPC title

• C07D403/14 Primary

  containing three or more hetero rings · CPC title

• A61K49/0058

  Antibodies · CPC title

• G01N33/582

  with fluorescent label · CPC title

• G01N33/533

  with fluorescent label · CPC title